中华妇产科杂志
中華婦產科雜誌
중화부산과잡지
CHINESE JOUNAL OF OBSTETRICS AND GYNECOLOGY
2014年
2期
114-119
,共6页
施秀%徐炜%孙莹%戴辉华%王秀丽
施秀%徐煒%孫瑩%戴輝華%王秀麗
시수%서위%손형%대휘화%왕수려
子宫内膜异位症%核受体共激活因子1%核受体共激活因子2%趋化因子CXCL12
子宮內膜異位癥%覈受體共激活因子1%覈受體共激活因子2%趨化因子CXCL12
자궁내막이위증%핵수체공격활인자1%핵수체공격활인자2%추화인자CXCL12
Endometriosis%Nuclear receptor coactivator 1%Nuclear receptor coactivator 2%Chemokine CXCL12
目的 探讨子宫内膜异位症(内异症)患者异位子宫内膜(异位内膜)基质细胞中甾体激素受体辅激活子(SRC)和基质细胞衍生因子1(SDF-1)的表达水平变化,了解不同类型SRC对雌二醇和孕酮调控的异位内膜基质细胞SDF-1表达的影响.方法 2010年5月至2012年10月,选择在南京医科大学第一附属医院妇科因卵巢内异症囊肿[美国生育学会修订的内异症分期标准(r-AFS)分期为Ⅲ～Ⅳ期]接受手术治疗,术后标本经病理检查证实为内异症的16例患者的异位内膜(增殖期9例,分泌期7例)及同期因非经期放置宫内节育器的无内异症病史且月经周期正常的健康妇女10例的正常内膜(增殖期5例,分泌期5例).所有患者术前3个月内均未服用过促性腺激素释放激素激动剂(GnRH-a)类药物.采用实时荧光定量PCR技术比较正常内膜和异位内膜在不同月经周期中SRC-1、SRC-2、SRC-3和SDF-1αmRNA表达水平.异位内膜基质细胞分离、培养、传代后,分别用含有10-8 moL/L的17β-雌二醇或10出mol/L的17β-雌二醇+10-6 mol/L孕酮的培养液继续培养.分别于培养24、48、72、96 h后收集上清液,采用ELISA法检测上清液中SDF-1α的蛋白表达水平.脂质体法分别介导SRC-1和SRC-2的小分子干扰RNA(siRNA)转染异位内膜基质细胞,转染后2d换用分别含有10-8mol/L的17β-雌二醇或10-8moL/L的17β-雌二醇+10-6mol/L孕酮的培养液继续培养3 d,ELISA法检测上清液中SDF-1α的蛋白表达水平.结果 (1)周期性变化:正常内膜中,SRC-1、SRC-2和SDF-1α的表达均具有周期性变化,增殖期mRNA的表达水平分别为5.6±1.2、3.8±1.1、2.7±0.5,分泌期的表达水平分别为2.6±1.0、2.1±1.0、1.6±0.5；分别比较,差异均有统计学意义(P＜0.05)；异位内膜缺乏周期性变化.(2)雌、孕激素培养不同时间后SDF-1α蛋白表达水平:17β-雌二醇培养异位内膜基质细胞48、72和96 h后,SDF-1α蛋白表达水平分别为(1 803±196)、(2 272±261)和(2 162±258) ng/L,17β-雌二醇+孕酮培养异位内膜基质细胞48、72和96 h后,SDF-1α蛋白表达水平分别为(1 307±150)、(1 518±301)和(1 550±144) ng/L.各时间点两者比较,差异均有统计学意义(P＜0.05).(3)沉默SRC基因对SDF-1α蛋白表达水平的影响:沉默异位内膜基质细胞中SRC-1后,17β-雌二醇培养72 h的异位内膜基质细胞SDF-1α蛋白的表达水平为(1 155±244) ng/L,较SRC-1沉默前的(2 313 ±357) ng/L下降50.04％,差异有统计学意义(P＜0.05).而沉默异位内膜基质细胞中SRC-2后,17β-雌二醇培养72 h的异位内膜基质细胞中SDF-1α蛋白表达水平为(1 958±324) ng/L,与沉默SRC-2前的(2 313 ±357) ng/L比较,差异无统计学意义(P＞0.05).沉默异位内膜基质细胞中SRC-2后,17β-雌二醇+孕酮培养72 h后的异位内膜基质细胞中SDF-1α蛋白表达水平为(2 051 ±380) ng/L,与沉默SRC-2前的(1 534±449) ng/L比较,差异有统计学意义(P＜0.05).结论 在甾体激素调控异位内膜基质细胞表达SDF-1α的作用中,SRC-1是雌激素的主要辅激活子,SRC-2是孕激素的主要辅激活子.
目的 探討子宮內膜異位癥(內異癥)患者異位子宮內膜(異位內膜)基質細胞中甾體激素受體輔激活子(SRC)和基質細胞衍生因子1(SDF-1)的錶達水平變化,瞭解不同類型SRC對雌二醇和孕酮調控的異位內膜基質細胞SDF-1錶達的影響.方法 2010年5月至2012年10月,選擇在南京醫科大學第一附屬醫院婦科因卵巢內異癥囊腫[美國生育學會脩訂的內異癥分期標準(r-AFS)分期為Ⅲ～Ⅳ期]接受手術治療,術後標本經病理檢查證實為內異癥的16例患者的異位內膜(增殖期9例,分泌期7例)及同期因非經期放置宮內節育器的無內異癥病史且月經週期正常的健康婦女10例的正常內膜(增殖期5例,分泌期5例).所有患者術前3箇月內均未服用過促性腺激素釋放激素激動劑(GnRH-a)類藥物.採用實時熒光定量PCR技術比較正常內膜和異位內膜在不同月經週期中SRC-1、SRC-2、SRC-3和SDF-1αmRNA錶達水平.異位內膜基質細胞分離、培養、傳代後,分彆用含有10-8 moL/L的17β-雌二醇或10齣mol/L的17β-雌二醇+10-6 mol/L孕酮的培養液繼續培養.分彆于培養24、48、72、96 h後收集上清液,採用ELISA法檢測上清液中SDF-1α的蛋白錶達水平.脂質體法分彆介導SRC-1和SRC-2的小分子榦擾RNA(siRNA)轉染異位內膜基質細胞,轉染後2d換用分彆含有10-8mol/L的17β-雌二醇或10-8moL/L的17β-雌二醇+10-6mol/L孕酮的培養液繼續培養3 d,ELISA法檢測上清液中SDF-1α的蛋白錶達水平.結果 (1)週期性變化:正常內膜中,SRC-1、SRC-2和SDF-1α的錶達均具有週期性變化,增殖期mRNA的錶達水平分彆為5.6±1.2、3.8±1.1、2.7±0.5,分泌期的錶達水平分彆為2.6±1.0、2.1±1.0、1.6±0.5；分彆比較,差異均有統計學意義(P＜0.05)；異位內膜缺乏週期性變化.(2)雌、孕激素培養不同時間後SDF-1α蛋白錶達水平:17β-雌二醇培養異位內膜基質細胞48、72和96 h後,SDF-1α蛋白錶達水平分彆為(1 803±196)、(2 272±261)和(2 162±258) ng/L,17β-雌二醇+孕酮培養異位內膜基質細胞48、72和96 h後,SDF-1α蛋白錶達水平分彆為(1 307±150)、(1 518±301)和(1 550±144) ng/L.各時間點兩者比較,差異均有統計學意義(P＜0.05).(3)沉默SRC基因對SDF-1α蛋白錶達水平的影響:沉默異位內膜基質細胞中SRC-1後,17β-雌二醇培養72 h的異位內膜基質細胞SDF-1α蛋白的錶達水平為(1 155±244) ng/L,較SRC-1沉默前的(2 313 ±357) ng/L下降50.04％,差異有統計學意義(P＜0.05).而沉默異位內膜基質細胞中SRC-2後,17β-雌二醇培養72 h的異位內膜基質細胞中SDF-1α蛋白錶達水平為(1 958±324) ng/L,與沉默SRC-2前的(2 313 ±357) ng/L比較,差異無統計學意義(P＞0.05).沉默異位內膜基質細胞中SRC-2後,17β-雌二醇+孕酮培養72 h後的異位內膜基質細胞中SDF-1α蛋白錶達水平為(2 051 ±380) ng/L,與沉默SRC-2前的(1 534±449) ng/L比較,差異有統計學意義(P＜0.05).結論 在甾體激素調控異位內膜基質細胞錶達SDF-1α的作用中,SRC-1是雌激素的主要輔激活子,SRC-2是孕激素的主要輔激活子.
목적 탐토자궁내막이위증(내이증)환자이위자궁내막(이위내막)기질세포중치체격소수체보격활자(SRC)화기질세포연생인자1(SDF-1)적표체수평변화,료해불동류형SRC대자이순화잉동조공적이위내막기질세포SDF-1표체적영향.방법 2010년5월지2012년10월,선택재남경의과대학제일부속의원부과인란소내이증낭종[미국생육학회수정적내이증분기표준(r-AFS)분기위Ⅲ～Ⅳ기]접수수술치료,술후표본경병리검사증실위내이증적16례환자적이위내막(증식기9례,분비기7례)급동기인비경기방치궁내절육기적무내이증병사차월경주기정상적건강부녀10례적정상내막(증식기5례,분비기5례).소유환자술전3개월내균미복용과촉성선격소석방격소격동제(GnRH-a)류약물.채용실시형광정량PCR기술비교정상내막화이위내막재불동월경주기중SRC-1、SRC-2、SRC-3화SDF-1αmRNA표체수평.이위내막기질세포분리、배양、전대후,분별용함유10-8 moL/L적17β-자이순혹10출mol/L적17β-자이순+10-6 mol/L잉동적배양액계속배양.분별우배양24、48、72、96 h후수집상청액,채용ELISA법검측상청액중SDF-1α적단백표체수평.지질체법분별개도SRC-1화SRC-2적소분자간우RNA(siRNA)전염이위내막기질세포,전염후2d환용분별함유10-8mol/L적17β-자이순혹10-8moL/L적17β-자이순+10-6mol/L잉동적배양액계속배양3 d,ELISA법검측상청액중SDF-1α적단백표체수평.결과 (1)주기성변화:정상내막중,SRC-1、SRC-2화SDF-1α적표체균구유주기성변화,증식기mRNA적표체수평분별위5.6±1.2、3.8±1.1、2.7±0.5,분비기적표체수평분별위2.6±1.0、2.1±1.0、1.6±0.5；분별비교,차이균유통계학의의(P＜0.05)；이위내막결핍주기성변화.(2)자、잉격소배양불동시간후SDF-1α단백표체수평:17β-자이순배양이위내막기질세포48、72화96 h후,SDF-1α단백표체수평분별위(1 803±196)、(2 272±261)화(2 162±258) ng/L,17β-자이순+잉동배양이위내막기질세포48、72화96 h후,SDF-1α단백표체수평분별위(1 307±150)、(1 518±301)화(1 550±144) ng/L.각시간점량자비교,차이균유통계학의의(P＜0.05).(3)침묵SRC기인대SDF-1α단백표체수평적영향:침묵이위내막기질세포중SRC-1후,17β-자이순배양72 h적이위내막기질세포SDF-1α단백적표체수평위(1 155±244) ng/L,교SRC-1침묵전적(2 313 ±357) ng/L하강50.04％,차이유통계학의의(P＜0.05).이침묵이위내막기질세포중SRC-2후,17β-자이순배양72 h적이위내막기질세포중SDF-1α단백표체수평위(1 958±324) ng/L,여침묵SRC-2전적(2 313 ±357) ng/L비교,차이무통계학의의(P＞0.05).침묵이위내막기질세포중SRC-2후,17β-자이순+잉동배양72 h후적이위내막기질세포중SDF-1α단백표체수평위(2 051 ±380) ng/L,여침묵SRC-2전적(1 534±449) ng/L비교,차이유통계학의의(P＜0.05).결론 재치체격소조공이위내막기질세포표체SDF-1α적작용중,SRC-1시자격소적주요보격활자,SRC-2시잉격소적주요보격활자.
Objectives To study the expression patterns of steroid receptor coactivators (SRC) and steroid-induced stromal cell-derived factor-1 (SDF-1) in endometriosis,and to explore the roles of SRC in the steroid-induced SDF-1 expression endometriosis.Methods From May 2010 to October 2012,16 endometriosis cases at stages Ⅲ or Ⅳ according to the revised American Society for Reproductive Medicine classification undergoing surgery in the First Affiliated Hospital to Nanjing Medical University were enrolled in this study.Their ectopic endometrium were from ovarian endometriomata which were identified pathologically with 9 cases at proliferative phase and 7 cases at secretory phase.The normal endometrium were acquired from the healthy women with normal menstrual cycle (n =10,proliferative phase =5,secretory phase =5).The mnRNA levels of SRC and SDF-1α during the menstrual cycle were detected by quantitative real-time polymerase chain reaction.Ectopic endometrium stromal cells were purified and cultured in medium containing 17β-estradiol (10-8mol/L) or 17β-estradiol (10-8 mol/L) + progesterone (10-6 mol/L).At 24,48,72 and 96 hours,the supernatants were collected to measure SDF-1α expression by ELISA.Ectopic endometrium stromal cells were transfected respectively with siRNA of SRC-1 and SRC-2 using lipofectamine.Two days after transfection,17β-estradiol (10-8 moL/L) or 17β-estradiol (10-8 mol/L) + progesterone (10-6 mol/L) were added into the media.On the third day after the steroid hormones treatment,the media were collected to quantify SDF-1α expression with ELISA.Results (1) Cyclical changes: the SRC-1,SRC-2 and SDF-1 α showed marked cyclic differences in normal endometrium (P ＜ 0.05).In proliferative phase and secretory phase,the SRC-1,SRC-2 and SDF-1 α were 5.6 ± 1.2,3.8 ± 1.1,2.7 ± 0.5 and 2.6 ± 1.0,2.1 ± 1.0,1.6-± 0.5,respectively.There was no periodic variation in the expression of SRC-1,SRC-2 and SDF-1α in ectopic endometrium throughout the menstrual cycle.(2) Steroid-induced SDF-1α expression in ectopic endometrium stromal cells: the 17β-estradiol-induced SDF-1α expression was (1 803 ± 196),(2 272 ± 261) and (2 162 ± 258) ng/L at 48,72 and 96 hours.At the same time points,the SDF-1α expression induced by 17β-estradiol and progesterone was (1 307 ± 150),(1 518 ± 301) and (1 550 ± 144) ng/L,respectively.There was significant difference between two groups (P ＜0.05).(3) The effects of SRC silencing on steroid hormones-induced SDF-1 α expression in ectopic endometrium stromal cells: the expression of 17β-estradiol-induced SDF-1α at 72 hours was significantly decreased from (2 313 ± 357) ng/L to (1 155 ± 244) ng/L after the silencing of SRC-1 (P ＜ 0.05).After the silencing of SRC-2,the 17β-estradiol-induced SDF-1 α at 72 hours was (1 958 ±324) ng/L.There was no significant difference compared with the before the silencing (P ＞ 0.05).The expression of SDF-1 α at 72 hours induced by 17β-estradiol + progesterone was (1 534 ± 449) ng/L and (2 051 ± 380) ng/L respectively before and after the silencing of SRC-2 and showed the significant difference (P ＜ 0.05).Conclusion During the expression of SDF-1 α regulated by steroids in ectopic endometrium cells,SRC-1 is the major coactivator of 17β-estradiol and SRC-2 is the major coactivator of progesterone.
